DESCRIPTION (applicant's description) We have recently shown that altering the expression of a small heat shock protein of carrot, Hsp 17.7, can significantly alter thermotolerance of both carrot and tomato (Zimmerman et al, 1999; Malik et al, 1999). This is the first demonstration in any plant system that alteration of the expression of a single gene can alternatively increase and decrease thermotolerance. Constitutive expression of Hsp 17.7 results in increased thermotolerance and also results in increased synthesis of all classes of heat shock proteins, especially under conditions of severe heat stress. In contrast, expression of an antisense version of the Hsp 17.7 gene leads to reduced thermotolerance and reduced synthesis of most heat shock proteins. These results show that even moderate changes in the amount of Hsp 17.7, either increasing or decreasing it, can cause a significant and pleiotropic response in the synthesis of other heat shock proteins and in thermotolerance, and suggest that Hsp 17.7 is a particularly important protein in the acquisition of thermotolerance. The goal of this proposal is to understand how Hsp 17.7 is effecting thermotolerance and why there is an apparent alteration in heat shock protein synthesis in cells that either over or under express it. We will use the transgenic cell lines we have generated to investigate the apparent link between Hsp 17.7 and the translation of other Hsps. Specifically we will address the following questions: (1) Is Hsp synthesis "primed" by the expression of Hsp 17.7 at room temperature in transgenic cells? (2) Is Hsp synthesis in the transgenic cells altered at the level of translation initiation or elongation? (3) Does lisp 17.7 directly interact with either the ribosome or nascent polypeptides of heat shocked cells, which could cause an enhancement of Hsp translation? We will characterize the Hsp 17.7 protein and analyze those properties that have been shown to be particularly important in the function of animal small Hsps, including phosphorylation and potential interaction with other cellular components such as the cytoskeleton, the ribosome, and other cytoplasmic proteins. These studies will result in a more complete understanding of the mechanism of action of this important plant small heat shock protein, and will provide insights into the thermotolerance of higher plants.